Colocasia esculenta L. schott corm mucilage: A selective COX-2 inhibitor for treatment of irritable bowel syndrome
Nesrin M Fayek1, Samar M Mouneir2, Azza Ramy Abdel Monem1, Samia M Abdelwahab1, Nebal D Eltanbouly1
1 Department of Pharmacognosy, Faculty of Pharmacy, Cairo University, Cairo, Egypt
2 Department of Pharmacology, Faculty of Veterinary Medicine, Cairo University, Cairo, Egypt
|Date of Submission||20-Oct-2020|
|Date of Decision||08-Jan-2021|
|Date of Acceptance||23-Mar-2021|
|Date of Web Publication||12-Jul-2021|
Azza Ramy Abdel Monem
Department of Pharmacognosy, Faculty of Pharmacy, Cairo University, Cairo 11562
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Therapeutic strategies used for the treatment of irritable bowel syndrome having many limitations due to their side effects; this necessitates searching for new substitutes with similar therapeutic results and limited side effects. In the present study, the anti-inflammatory activity of the mucilage of Colocasia esculenta L. Schott var. typical corm cultivated in Egypt was explored both in vivo and in vitro. Materials and Methods: In vivo anti-inflammatory activity was evaluated based on histopathological examination, determination of ulcer area and ulcer index, and measurement of inflammatory mediators, namely, myeloperoxidase, malondialdehyde, nitric oxide, and tumor necrosis factor-α, in acetic acid-induced ulcerated rat colon, comparing to Prednisolone as a reference drug. In vitro evaluation of the anti-inflammatory activity of the tested mucilage was carried out by measuring its cyclooxygenase (COX-1/COX-2) and 5-lipoxygenase inhibitory activity comparing to Celecoxib and Zileuton as reference drugs, respectively. Results: Pretreatment with the mucilage improved the histopathological features of the rats ulcerated colon and decreased the ulcer area and the ulcer index in a dose-dependent manner. The mucilage improved all the tested inflammatory mediators. In addition, it has a potent COX-2 inhibitory activity with lower effect on COX-1 and 5-lipooxygenase. Conclusion: The obtained results support the use of C. esculenta corm mucilage as an alternative for the treatment of inflammatory bowel disease with minimum gastrointestinal side effects.
Keywords: Anti-inflammatory activity, Colocasia esculenta, mucilage
|How to cite this article:|
Fayek NM, Mouneir SM, Abdel Monem AR, Abdelwahab SM, Eltanbouly ND. Colocasia esculenta L. schott corm mucilage: A selective COX-2 inhibitor for treatment of irritable bowel syndrome. Phcog Mag 2021;17:387-90
|How to cite this URL:|
Fayek NM, Mouneir SM, Abdel Monem AR, Abdelwahab SM, Eltanbouly ND. Colocasia esculenta L. schott corm mucilage: A selective COX-2 inhibitor for treatment of irritable bowel syndrome. Phcog Mag [serial online] 2021 [cited 2021 Aug 3];17:387-90. Available from: http://www.phcog.com/text.asp?2021/17/74/387/321253
- Colocasia esculenta corm mucilage improved the histopathological features in acetic acid induced ulcerated rat colon and decreased the ulcer area and ulcer index in a dose dependent manner. The mucilage improved all the tested inflammatory mediators viz. myeloperoxidase, malondialdehyde, nitric oxide and tumor necrosis factor-α, comparing to Prednisolone. The mucilage showed a potent cyclooxygenase-2 inhibitory activity with lower effect on cyclooxygenase-1 and 5-lipooxygenase comparing to Celecoxib and Zileuton, respectively.
Abbreviation list: COX: cyclooxygenase; IBD: inflammatory bowel disease; LOX: lipoxygenase; MDA: malondialdehyde; MPO: myeloperoxidase; NO: nitric oxide; p.o.: per oral; pg/mg: picograms/mg; TNF-α: tumor necrosis factor-α; g: gram; h: hour; mg/kg/day: milligram/kilogram/day; IC50: Half-maximal inhibitory concentration; mg/kg b.wt: milligram/kilogram/body weight; μg/mL: microgram/milliliter
| Introduction|| |
Inflammatory diseases of the intestinal tract (inflammatory bowel disease [IBD]) include ulcerative colitis and Crohn's disease. Ulcerative colitis is limited to the colon and/or rectum and affects only the inner lining of the intestine, while Crohn's disease can affect any part of the gut from mouth to anus as skip lesions and affect the whole thickness of the bowel wall. Nonsteroidal anti-inflammatory drugs, worldwide, are commonly used for excellent anti-inflammatory effect, but with limited use as a result of their severe side effects such as gastrointestinal ulceration, perforation, obstruction, and bleeding. This suggests that seeking for other drugs harboring similar therapeutic results and limited side effects are urgently necessary. Several reports describe a significant anti-inflammatory activity for plants rich in mucilage, for example, Trigonella foenum-graecum and Aloe littoralis. Colocasia esculenta L. Schott corm is rich with mucilage, up to 22.3%, and it is used in Egypt as a food. Although previous studies proved that C. esculenta L. Schott leaves have anti-inflammatory activity,, no study has been performed to evaluate the anti-inflammatory activity of C. esculenta L. Schott corm mucilage. Consequently, this study was carried out to evaluate the in vivo and in vitro anti-inflammatory activity of the mucilage of C. esculenta corms cultivated in Egypt.
| Materials and Methods|| |
The fresh C. esculenta L. Schott var. typical corms were collected in 1st week of January 2017 from the experimental field in Faculty of Agriculture. The plant was identified by Prof. Dr. Hassan Ali Hassan, Department of vegetable crops, Faculty of Agriculture, Cairo University, Giza, Egypt.
Preparation of mucilage
Preparation of mucilage was done following the method described by Sarkar et al.
Drugs and chemicals
Acetic acid was purchased from Merck, Darmstadt, Germany. Prednisolone, Celecoxib and Zileuton, as an anti-inflammatory reference drugs, were obtained from Chemical Industries Development Co., Giza, Egypt.
Three-month-old male and female Wistar strain rats (130–150 g) were purchased from laboratory animal house of Faculty of Veterinary Medicine, Cairo University. The animal room was maintained at 22°C–24°C and a lighting regimen of 12 h light/12 h dark. Animals had free access to standard house chow and water ad libitum. All experimental procedures were performed according to the ethical roles of Faculty of Veterinary Medicine, Cairo University.
In vivo evaluation of the anti-inflammatory effect
Induction of ulcerative colitis
Experimental ulceration in colon tissue was done according to the method described by Al-Rejaie et al. 2013.
Twenty-five animals were divided into five groups (five animals in each) as follow: Group I: Normal animals; received 2 mg/kg/day of distilled water. Group II: Acetic acid control animals; received 2 mL of 4% acetic acid solution intrarectally on the 8th day. Group III: Prednisolone standard treated animals; received prednisolone (2 mg/kg, p.o., for 3 days) and acetic acid (2 mL of 4% solution, once on the 8th day, intrarectally). Prednisolone and acetic acid treatment were started on the same day. Group IV: Mucilage (10 mg/kg) treated animals; received 7 days pretreatment with 10 mg/kg of mucilage, p. o. and 2 mL of 4% acetic acid solution, intrarectally on the 8th day. Drug treatment was continued till 11th day. Group V: Mucilage (20 mg/kg)-treated animals; received 7 days pretreatment with 20 mg/kg of mucilage, p.o., and 2 mL of 4% acetic acid solution, intrarectally on the 8th day. Drug treatment was continued till the 11th day.
On the 11th day, animals were euthanized by over dose of chloroform anesthesia and their colons were collected for examination and preparation of tissue homogenate.
Autopsy samples were taken from the colon of rats in the different animal groups and prepared for microscopic examination following the method of Banchroft et al.
Determination of ulcer area and ulcer index was done according to Kandhare et al.
Samples from the colon were stored immediately at −80°C till analysis. Tissue samples were homogenized in 10 mmol Tris–HCl buffer (pH 7.1), and the homogenate was utilized for the measurement of inflammatory mediators. Myeloperoxidase (MPO) and malondialdehyde (MDA) detection kits were purchased from Hycult biotech and Sigma-Aldrich Co., respectively, while nitric oxide (NO) and tumor necrosis factor-α (TNF-α) detection kits were purchased from Thermo Fisher Scientific and RayBiotech, Inc., respectively.
In vitro evaluation of the anti-inflammatory effect
Cyclooxygenase-1/cyclooxygenase-2 inhibition assay
The inhibitory cyclooxygenase (COX) activity of the tested mucilage and the reference drug (Celecoxib) was assayed using Cayman colorimetric COX (ovine) inhibitor screening assay kit (Catalog No. 760111), purchased from Cayman chemicals, MI, USA and IBL International GmbH, Hamburg, Germany according to the manufacturer's instructions.
5-lipoxygenase inhibition assay
It was carried out on the tested mucilage and the reference drug (Zileuton) using CM760700 kit, purchased from Cayman chemicals, MI, USA and IBL International GmbH, Hamburg, Germany, according to the manufacturer's instructions.
Data of in vivo anti-inflammatory activity were expressed as the mean ± standard deviation, determination was carried out in triplicates. The statistical significance of differences between the mean values was analyzed by one-way analysis of variance followed by posthoc test and least-significant difference, using SPSS software (SPSS INnc., Chicago, USA). A value of P < 0.05 was considered statistically significant for analysis.
The IC50 values of in vitro anti-inflammatory activity and anticancer activity were calculated according to the equation for the Boltzmann sigmoidal concentration response curve using non-linear regression fitting models (Graph Pad, Prism Version 5 (GraphPad Software, San Diego, California)).
| Results|| |
In vivo anti-inflammatory activity
The histopathological investigation as presented in [Figure 1], showing no alteration in the control negative group [Group I and [Figure 1]a] with normal histological structure of the mucosa with glandular structure and lamina propria and the underlying submucosa, muscularis and serosa. In the control positive group [Group II and [Figure 1]b], focal ulceration and necrosis with inflammatory cells infiltration were detected in the mucosa, submucosa, and serosal layers. In the standard (Prednisolone) group [Group III, [Figure 1]c], the mucosa was intact, while the underlying submucosa showed inflammatory cells infiltration. In the group treated with 10 mg/kg b.wt of mucilage [Group IV, [Figure 1]d], focal ulceration and necrosis were seen, while the underlying submucosa had inflammatory cells infiltration and hemorrhages. In higher dose of the mucilage [20 mg/kg. b.wt, Group V, [Figure 1]e], edema with inflammatory cells infiltration were seen and congested blood vessels were recorded in the lamina propria of the mucosa as well as in the submucosa.
|Figure 1: Histopathological changes of (a) control negative group; (b) control positive group; (c) standard (prednisolone) group; (d) treated group with 10 mg/kg. b.wt of mucilage; (e) treated group with 20 mg/kg b.wt of mucilage (X = 40)|
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Effect of colocasia mucilage on ulcer area and ulcer index
The mean ulcer area and ulcer index of acetic acid control group were (27.8 ± 1.92) mm2 and (55.6 ± 1.94), respectively, indicating high ulcerogenic effect of acetic acid. Pretreatment of Colocasia mucilage (10 and 20 mg/kg, p.o.) for 7 days decreased the ulcer area ([20 ± 2.54] and [11.6 ± 1.51] mm2, respectively) and the ulcer index ([22.8 ± 1.92] and [18.6 ± 2.40], respectively) as compared to acetic acid control group in a dose-dependent manner [Table 1].
|Table 1: Effect of Colocasia mucilage on the ulcer index in acetic acid-induced inflammatory bowel disease in rats|
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Inflammatory and oxidative alternations of the colonic mucosa
Seven-day pretreatment with mucilage significantly reduced the acetic acid-induced colonic mucus content and prevented oxidative and inflammatory response in a dose-dependent manner [Table 2] and [Figure 2]. Group V, receiving the higher dose of the mucilage (20 mg/kg), showed higher decrease (82.7%, 69.9%, 73.5%, and 86.1%) in the MPO, MDA, NO, and TNF-α levels, respectively, in comparison to prednisolone (92.1%, 85.6%, 88.3%, and 93.6%, respectively). Meanwhile, the second mucilage-treated group (Group IV; 10 mg/kg) showed lower decrease in the same mentioned parameters (67.4%, 56.1%, 61.6%, and 72.9%, respectively).
|Table 2: Effect of Colocasia mucilage on the myeloperoxidase, malondialdehyde, nitric oxide, and tumor necrosis factor-α activity in acetic acid-induced colitis in rats|
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|Figure 2: Effect of Colocasia esculenta L. Schott corm mucilage on myeloperoxidase, malondialdehyde, nitric oxide and tumor necrosis factor-α activity in acetic acid-induced rat colitis comparing to reference drugs|
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In vitro anti-inflammatory activity
Cyclooxygenase-1/cyclooxygenase-2 inhibition assay
The obtained results [Table 3] and [Figure 3] revealed that C. esculenta mucilage has a potent COX-2 inhibitory activity (IC50 of 0.027 μg/mL) more than the standard drug, Celecoxib (IC50 of 0.034 μg/mL), with lower effect on COX-1 (IC50 of 1.74 μg/mL) upon comparing to Celecoxib (IC50 of 1.21 μg/mL).
|Table 3: Effect of Colocasia esculenta mucilage (IC50 values) on cyclooxygenase-1, cyclooxygenase-2, and 5-lipoxygenase activity comparing to standard drugs|
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|Figure 3: IC50 of the Colocasia esculenta L. Schott corm mucilage on COX-1, COX-2 and 5-LOX activity in acetic acid-induced rat colitis comparing to reference drugs|
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5-Lipoxygenase inhibition assay
The mucilage showed lower 5-lipoxygenase (5-LOX) inhibitory activity (IC50 of 0.57 μg/mL) than the standard drug, Zileuton (IC50 of 0.33 μg/mL) [Table 3] and [Figure 3].
| Discussion|| |
Intrarectal administration of acetic acid causes protonation and migration of acetic acid molecule into colonic microflora, leading to epithelial denudation followed by infiltration of neutrophils into colonic tissue which indicates inflammatory conditions. The histopathological examination highlighted the reducing effect of the mucilage on the damage scores of acetic acid comparing to prednisolone as a standard drug. The MPO and TNF-α activities were the main parameters for evaluating the degree of acetic acid-induced colonic inflammation in rats, while MDA and NO content were the prominent parameters for evaluating the oxidative stress of the colonic mucosa. Compared with the normal group (Group I), all these parameters were significantly increased by acetic acid (P < 0.05) in the colonic mucosa of the model group (Group II). After the model rats were treated with mucilage (10, 20 mg/kg; Group IV and V) or prednisolone (2 mg/kg; Group III), these elevated parameters were significantly improved in a dose-dependent manner (P < 0.05). Accordingly, the therapeutic dose protocol of 20 mg/kg of the mucilage (Group V) can be considered as an effective drug in the treatment of this rat model of colitis. In addition, this natural mucilage overcomes the side effects of prednisolone-like anxiety, blurred vision, nervousness, depression, and restlessness and hence, mucilage can be prescribed for geriatric patients. The mucilage forms a protective viscous layer that could attenuate irritation, independently from the origin (mechanic, thermic, chemical, etc.).
A promising approach in the treatment of inflammatory diseases includes the inhibition of COX and LOX pathways. COX is a prostaglandin endoperoxide synthase enzyme. In mammalian cells, COX exists in at least two isoforms COX-1 (expressed in all cell types and regulated as a house keeping enzyme for various physiological functions) and COX-2 (expressed during tissue damage or inflammation). 5-LOX pathway is the major source of potent pro-inflammatory leukotriene's.
Nonsteroidal anti-inflammatory drugs possess excellent anti-inflammatory effect through inhibition of COX-2, but with limited use as a result of its severe side effects on gastrointestinal tract through inhibition of COX-1. Nevertheless, the anti-inflammatory agents having selective COX-2 inhibition but less reactive toward COX-1 are appreciated as novel anti-inflammatory agents in the mainstream of anti-inflammatory research. The promising anti-inflammatory activity of C. esculenta corm mucilage compared to Celecoxib makes it good candidates for further optimization and development of potent and safe anti-inflammatory agents having selective COX-2 inhibition with minimum gastrointestinal side effects.
The findings of the present study strongly suggested that the freeze-dried mucilage of Egyptian cultivated C. esculenta var. typical corm could serve as an alternative modality for the treatment of IBD and may pose promising outcomes for future clinical usage of mucilage as a natural nontoxic effective supplement in IBD. However, further fractionation of the mucilage needs to be carried out to determine the bioactive moieties particularly responsible for these activities.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3]
[Table 1], [Table 2], [Table 3]